1. Field of the Invention
The present invention is directed to an in vitro immunodeficiency virus neutralization assay.
2. Description of Related Art
In order to develop useful vaccines, drugs and other therapeutic agents for treating and/or preventing human immunodeficiency virus (HIV) infections in humans, it is very useful to evaluate these vaccines, drugs and therapeutic agents in a cell culture (in vitro) system to determine their ability to inhibit or interfere with viral replication. In this respect, there has been a considerable amount of work with antibodies which are reactive with antigens present in or on HIV. These antibodies may arise as a result of HIV infection and, thus, are diagnostic of this infection or these antibodies may be elicited by exposure to candidate HIV vaccines and, thus, may predict protection from future infection or prevention of disease progression. In the past, these antibodies have usually been evaluated in an in vitro cell culture system which utilizes a continuous cell line, Potts et al, Virology, 197, 415-419(1993), White-Scharf et al, Virology, 192, 197-206(1993), Ho et al, J. Virol. 61, 2024-2028(1987), Javaherian et al, PNAS USA, 89, 1418-1422(1992), Broliden et al, PNAS USA, 89, 461-465(1992), Welch et al, J. Clin. Microbiol., 30, 1424-1427(1992) and Skinner et al, J. Virol., 62, 4195-4200(1988). In these most commonly used known systems, the antibody sample to be tested and the virus are mixed with each other and incubated for a period of time (usually one hour or more) and the antibody/virus mixture is then added to the cells. However, a continuous cell culture system is not an ideal system for predicting the effectiveness of an antibody in preventing HIV replication in vivo since there are numerous biological and physiological differences between the highly selected and potentially transformed cells which make up a continuous cell line and the normal cells which are present in the human body.
There have been several reports for measuring the ability of anti-HIV antibodies to neutralize laboratory strains and clinical isolates of HIV using primary human lymphocytes, also referred to as peripheral blood mononuclear cells (PBMC), Kliks et al, PNAS USA 90, 11518-11522(1993), Albert et al, Aids Res. Human Retroviruses, 9, 501-514(1993), Albert et al, AIDS 4, 107-112(1990), Cheng-Mayer et al, PNAS USA, 85, 2815-2819(1988) and Homsy et al, J. Virol., 64, 1437-1440(1990). Laboratory strains of HIV are different from primary clinical isolates of HIV in that they have been adapted via mutation to replicate in continuous lymphocytic cell lines. There are limited published reports of neutralization of clinical HIV isolates in peripheral blood mononuclear cells. In all these assays, the laboratory strains and clinical isolates of HIV and antibody samples were preincubated for a period of time prior to addition to the target peripheral blood mononuclear cells. In addition, the clinical HIV isolates used in all studies except one, Albert et al, AIDS, 4, 107-112(1990) had been passaged in the laboratory since initial isolation in 1984 and 1985, Levy and Shimabukuro, J. Infect. Dis., 152, 734-738(1985), Levy et al, Science, 225, 840-842(1984), and Levy et al, Lancet, 11, 586-588(1985) and, thus, cannot be considered to be primary isolates of HIV.